P53 AND PROTEIN-KINASE-C INDEPENDENT INDUCTION OF GROWTH ARREST AND APOPTOSIS BY BRYOSTATIN-1 IN A HIGHLY METASTATIC MAMMARY EPITHELIAL-CELL LINE - IN-VITRO VERSUS IN-VIVO ACTIVITY

We have evaluated the effects of bryostatin I on growth of a highly ma lignant p53-null mouse mammary tumor line, 4T1, and the mechanism by w hich bryostatin I inhibits in vitro growth and in vivo development of tumor and metastases from the orthotopic site. Bryostatin I at 20-400 nM concentrations inhibits growth of 4T1 cells by similar to 60% in tw o-day cultures. Inhibition of growth is associated with an increase in the number of cells undergoing apoptosis with concomitant elevation i n the steady state levels of bar protein and drop in bcl-2 levels. The cytotoxic effect of bryostatin 1 on 4T1 cells occurs independently of p53, since there was no evidence of p53-mediated transcriptional acti vity in 4T1 cells following treatment with bryostatin 1. 4T1 cells res pond in vivo to bryostatin 1 therapy (75 mu g/kg body weight). Intrape ritoneal administration of bryostatin 1 inhibits both primary and seco ndary tumor growth by similar to 50%. However, although bryostatin 1 h as a remarkable capacity to slow tumor growth and progression, it is u nable to completely eradicate tumor growth and progression due to in v ivo development of tumor resistance to bryostatin 1. Levels and cellul ar distribution of PKC alpha and delta do not correlate with the growt h inhibitory effects of bryostatin 1 on 4T1 cells; however, reduction in cytosolic PKC alpha and delta without associated increase in membra ne compartment appear to correlate with bryostatin-resistance. Our res ults suggest that the therapeutic effects of bryostatin 1 in our syste m do not involve alterations in levels and distribution of PKC but rat her a direct upregulation of bax/bcl-2 ratios that is independent of p 53.